GPR39-1b, the 5-transmembrane isoform of GPR39 interacts with neurotensin receptor NTSR1 and modifies its function

Abstract

G-protein-coupled receptor 39 (GPR39), a member of the ghrelin receptor family, has a full-length isoform GPR39-1a and a truncated isoform GPR39-1b. While GPR39-1a was clarified as a receptor of Zn²⁺, the characteristic property of GPR39-1b remains unknown. Therefore, in this study, the molecular functions of GPR39-1b were explored in cell culture. In contrast to GPR39-1a, GPR39-1b showed no response to Zn²⁺ stimulation in calcium mobilization assays, suggesting that GPR39-1b is not a functional receptor of Zn²⁺. To understand the signaling interaction of GPR39-1b, we investigated the dimerization between the isoforms, and conducted bioluminescence resonance energy transfer (BRET²) assays. The results indicated that GPR39-1b homodimerized, but did not heterodimerize with GPR39-1a. We subsequently attempted to search the heterodimeric counterparts of GPR39-1b. Neurotensin receptor 1 (NTSR1) was also targeted as a GPR39-1b interacting partner because of its highly conserved amino acid sequence and mRNA localization, which was similar to GPR39-1b. BRET² assays demonstrated that GPR39-1b heterodimerized with NTSR1. To examine the effect of GPR39-1b on NTRS1-mediated cAMP/PKA signaling, we used the cAMP responsive element-luciferase assays and observed that GPR39-1b attenuated neurotensin-induced NTSR1 signaling. Taken together, our results provided a novel regulatory mechanism for GPR39-1b in NTRS1 signaling.

• BRET
• G-protein-coupled receptor
• GPR39
• heterodimerization
• neurotensin
• NTSR1

Acknowledgements

The authors thank Dr Emiko Sugikawa, Dr Mitsuhiro Kikyo, Moeko Kanai, Yasufumi Fukano, Dr Kenji Arakawa, Dr Masaaki Nagasaki and Dr Akiyoshi Fukamizu for scientific suggestions and technical instructions.